Chloroquine transfection protocol

This procedure can be modified for alternative packaging cell lines or transfection reagents. Once produced, lentivirus can be used for a variety of downstream applications such as stable-cell line generation.

When performing a transient transfection, you can choose between a standard or reverse transfectionprotocol. In a standard transfectionprotocol, the cells are plated on day 1, transfected on day 2 and assayed on day 3 or 4. In a reverse transfectionprotocol, cells are added directly to a plate containing the transfection reagent/DNA mix and. Prior to adding complex, remove 1 mL of medium from each well, leaving a total of 1 mL in each well. Add 500 μL of lipid–DNA complex to each well, taking care to dispense liquid against the well wall to avoid disrupting cells. Gently agitate plate to evenly distribute. Incubate plate for 6 hours at 37°C, 5% CO 2. This protocol is modified from the Addgene website protocol Materials. Transfection Procedure. The morning of the transfection day, replace the media with fresh DMEM without PSG and containing 10 uL of 25 mM chloroquine optional. Wait ~5h before going onto the next step.

Test a variety of brands and lots of FBS to find one suitable with your protocols. Last Upload: August 26, 2016 *Pro-Tip* Different brands and lots of FBS can promote or inhibit transfection.

Chloroquine is an inhibitor of the lysosomal degradation of the DNA which is taken up by the cells, so as leelee said, transfection should have been successful, albeit at a slightly lower level than if you would have added the chloroquine. In my normal transfections, I never add chloroquine and get high efficiencies nevertheless. Chloroquine treatment of rodent cells during the first hours of polyoma DNA transfection increase the fraction of cells expressing viral functions. The effect has been observed after DNA absorption using both the DEAE-dextran and calcium phosphate coprecipitation methods. Of the transfection complex I forgot to treat the cells with This protocol describes a general method for transfecting mammalian cells using Dissolve 0.129 g of chloroquine diphosphate salt into 10 mL of sterile 26, 2016 Use this protocol to generate lentivirus. 293T cells using a polyethyenimine

Hydroxychloroquine is used to treat or prevent malaria, a disease caused by parasites that enter the body through the bite of a mosquito. Hydroxychloroquine is also used to treat symptoms of rheumatoid arthritis and discoid or systemic lupus erythematosus.

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Hydroxychloroquine Side-effects, uses, time to work However, in some people hydroxychloroquine can cause skin rashes, especially those made worse by sunlight. feeling sick nausea or indigestion. diarrhoea. headaches. bleaching of the hair or mild hair loss. tinnitus ringing in the ears. visual problems.